Identification of human miRNA precursors that resemble box C/D snoRNAs

Published online 18 January 2011 Nucleic Acids Research, 2011, Vol. 39, No. 9 3879–3891 doi:10.1093/nar/gkq1355 Identification of human miRNA precursors that resemble box C/D snoRNAs Motoharu Ono1, Michelle S. Scott2, Kayo Yamada1, Fabio Avolio1, Geoffrey J. Barton2 and Angus I. Lamond1,* 1 Wellcome Trust Centre for Gene Regulation and Expression and 2Division of Biological Chemistry and Drug Discovery, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, UK Received October 29, 2010; Revised December 22, 2010; Accepted December 23, 2010 ABSTRACT There are two main classes of small nucleolar RNAs (snoRNAs): the box C/D snoRNAs and the box H/ ACA snoRNAs that function as guide RNAs to direct sequence-specific modification of rRNA precursors and other nucleolar RNA targets. A previous computational and biochemical analysis revealed a possible evolutionary relationship between miRNA precursors and some box H/ACA snoRNAs. Here, we investigate a similar evolutionary relationship between a subset of miRNA precursors and box C/ D snoRNAs. Computational analyses identified 84 intronic miRNAs that are encoded within either box C/D snoRNAs, or in precursors showing similarity to box C/D snoRNAs. Predictions of the folded structures of these box C/D snoRNA-like miRNA precursors resemble the structures of known box C/D snoRNAs, with the boxes C and D often in close proximity in the folded molecule. All five box C/D snoRNA-like miRNA precursors tested (miR-27b, miR-16-1, mir-28, miR-31 and let-7g) bind to fibrillarin, a specific protein component of functional box C/D snoRNP complexes. The data suggest that a subset of small regulatory RNAs may have evolved from box C/D snoRNAs. INTRODUCTION Micro RNAs (miRNAs) are a family of short regulatory RNAs that post-transcriptionally regulate gene expression. In mammals, miRNAs have been found to perform their regulatory function mainly by translation inhibition of protein coding transcripts through base pairing to specific target sequences in the 30 -untranslated regions (UTRs) (1). While a subset of miRNAs are encoded in independent transcription units, many miRNAs are encoded in introns of protein-coding genes and are co-expressed with these host genes (2–4). Mature miRNAs are small RNAs of 22 nt in length that are processed out of 70 nt-long hairpin structures (called pre-miRNAs) (5). The canonical miRNA biogenesis pathway involves either excision of the miRNA precursors from the introns of their host gene transcripts or transcription from independent units, both followed by processing by the microprocessor complex in the nucleus, export to the cytoplasm and further processing by a dicer-containing complex (2–4). However, recent reports have identified several different non-canonical miRNA processing pathways (6–9). In particular, several groups have recently reported miRNAs and miRNA-like molecules derived from small nucleolar RNAs (snoRNAs), some of which have been immunoprecipitated with Ago proteins, functional protein interactors of mature miRNAs (7,8,10,11). snoRNAs are a family of conserved nuclear RNAs concentrated in nucleoli where they either function in the modification of ribosomal RNA (rRNA) or participate in the processing of rRNA during ribosome subunit synthesis (12–15). Most snoRNAs have been found to be encoded in the introns of protein-coding genes (16). snoRNAs are processed out of these introns and carry out their function in complex with specific protein interactors, forming ribonucleoprotein complexes referred to as snoRNPs. Two main classes of snoRNAs have been identified: the box C/D snoRNAs and the box H/ACA snoRNAs, both of which serve as guide RNAs complementary to specific target sequences mainly in rRNA precursors. Box C/D snoRNAs catalyse 20 -Oribose methylation and box H/ACA snoRNAs guide psuedouridine modifications. *To whom correspondence should be addressed. Tel: +44 (0)1382 385473; Fax: +44 (0)1382 388072; Email: Present address: Fabio Avolio, Molecular Biotechnology Centre, University of Turin, Via Nizza 52, 10126 Torino, Italy. The authors wish it to be known that, in their opinion, the first two authors should be regarded as joint First Authors. ß The Author(s) 2011. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. 3880 Nucleic Acids Research, 2011, Vol. 39, No. 9 Box C/D snoRNAs are characterized by the presence of conserved C and D sequence boxes that typically come into contact in the folded molecule and serve as a binding site for box C/D snoRNP proteins, including NOP56, NOP58, 15.5K and the highly conserved protein fibrillarin, which carries out the specific 20 -O-methylation. The guide sequence with complementarity to the target is located immediately 50 to the box D or D0 region (e.g. see Figure 1). Several box C/D snoRNAs and a subset of their target sites in rRNA are conserved from yeast through to mammalian cells. However, numerous orphan box C/D snoRNAs have been identified that do not encode a region of complementarity to rRNA (17). Through systematic investigation of human miRNAs, we have recently reported that a subset of miRNA precursors have box H/ACA snoRNA features, both in their primary and secondary structure as well as in the genomic region in which they are encoded. Five of these miRNA precursors show functional box H/ACA snoRNA characteristics by binding to dyskerin, a conserved protein component of the box H/ACA snoRNPs. This led us to propose an evolutionary relationship between miRNAs and snoRNAs, in which a subset of snoRNAs would have evolved to serve as precursors of miRNAs (10). snoRNAs have been characterized as mobile genetic elements with the capacity to copy themselves to other genomic locations (18,19), thus providing large numbers of potential miRNA-like precursors. Such copies of snoRNAs have been coined snoRTs [snoRNA retroposons, by Weber (18)]. Recent reports provide further complementary evidence supporting this hypothesis, including the discovery of nucleolar miRNAs (20) and the description of small miRNA size fragments derived from snoRNAs (11,21). In addition, we have recently described a family of closely related snoRNAs, the HBII-180s, which show typical features of box C/D snoRNAs but also contain a region of 20 nt with almost perfect complementarity to endogenous pre-mRNA sequences, termed the M-box (22). Although the detailed mechanism and relationship between the M-box sequences and the endogenous target RNAs to which they are complementary has not yet been characterized, we demonstrated that by altering the M-box region to make it complementary to selected target genes it is possible to suppress target gene mRNA and protein le (...truncated)